The Amyloid Precursor Protein (APP) and its metabolism play fundamental roles in the pathophysiology of Alzheimer's disease (AD). Processing of APP along the amyloidogenic pathway results in the generation of the neurotoxic Aβ peptide, while cleavage along the non-amyloidogenic pathway generates sAPPα, which is known to exhibit neuroprotective and neurotrophic properties. With regard to these diametral functions of APP cleavage products it is supposed that not only the pathological accumulation of Aβ but also the loss of the physiological function of sAPPα could contribute to the neurodegenerative effects and the cognitive decline in patients with Alzheimer`s disease
The aim of the following study was to investigate the biological properties of the neuroprotective peptide sAPPα under different Alzheimer-relevant stress conditions. In the first part of the thesis proteasomal stress, which is closely associated with AD, was induced by the application of the proteasomal inhibitor Epoxomicin. To investigate the properties of sAPPα, conditioned media of APP-overexpressing HEK293 cells or purified recombinant sAPPα from different expression systems were used. Preincubation with sAPPα antagonized stress-triggered cell death in PC12 cells. Morphological analysis of fragmented nuclei, the uptake of propidium iodide and the analysis of effector-caspase activity confirmed the protective abilities of sAPPα but not of sAPPβ, which differs only in 17 amino acids.
The second stress model was established by the application of Thapsigargin, which is known to disturb the intracellular Ca2+-homoeostasis by inhibiting sarco/endoplasmic Ca2+-ATPases. This disruption is a common feature in brain ageing and the pathophysiology of AD, and was employed to induce apoptosis in the present study. Preincubation with sAPPα again showed the ability to antagonize this effect both in PC12 cells and in hippocampal neurons. The protective effect of sAPPα is partially mediated by the inhibition of the pro-apoptotic JNK/c-Jun signalling pathway and the activation of the anti-apoptotic PI3K/Akt signalling pathway, which was shown in both stress models.
One possible target for pharmacological intervention is the proteolytic processing of APP. Accordingly, the stimulation of the non-amyloidogenic processing pathway resulting in a higher production of the neuroprotective fragment sAPPα could antagonize the neurotoxic effects of Aβ. In the present study omega-3 fatty acids, e. g. docosahexaenoic acid (DHA), were used to modulate membrane fluidity of APP overexpressing HEK293 cells. The preincubation of the HEK-APP cells with DHA was associated with an increase in membrane fluidity closely correlating with an enhanced secretion of sAPPα. This observed increased non-amyloidogenic APP processing due to the DHA treatment was associated with a protection against apoptosis induced by the disturbance of the sensitive Ca2+-homoeostasis. Finally it was possible to confirm an essential role of sAPPα in mediating the DHA-dependent protective effect.